A Sensitive and Ara h 2 Specific Competitive ELISA for the Detection of Peanut in Processed Foods

Introduction: The food industry relies heavily on ELISA-based methods for detecting peanut in food matrices. Although commercial peanut ELISA kits perform quite well with many food matrices, most of them do not perform as well with highly heat processed foods, mostly because they predominantly detect the heat sensitive peanut allergen Ara h 3.

Purpose: The objective of this study was to develop a more robust and sensitive peanut ELISA targeting the heat stable peanut allergen Ara h 2.

Methods: Peanut Ara h 2 was purified from light roast peanut flour using column chromatography techniques. Polyclonal antibodies raised in rabbits against purified Ara h 2 were used to develop a competitive inhibition ELISA targeting Ara h 2. The performance of the developed ELISA in recovering peanut from a solid food matrix (pastry squares) subjected to different processing conditions was compared with that of two commercial peanut ELISA kits.

Results: The developed ELISA was highly sensitive with a detection range of 40 to 400 ng/ml of Ara h 2. The limit of detection and IC₅₀were 27±8 ng/ml and 134±32 ng/ml of Ara h 2, respectively. The Ara h 2 ELISA performed significantly better (P<0.05) than both the Neogen Veratox and Morinaga Peanut ELISA methods in recovering peanut from fried, baked and high pressure processed pastry squares incurred with peanut. With samples subjected to more severe processing, the performance of the developed ELISA was comparable (P>0.05) to the Morinaga ELISA. Among 45 food ingredients tested for cross reactivity with the rabbit antisera, none showed any cross reactivity although minor matrix interference was observed with cloves, cocoa powder, mustard, and pistachios.

Significance: The results indicate that the developed ELISA, targeting a heat stable peanut allergen, shows higher sensitivity than some of the currently available ELISAs in detecting peanut from processed food matrices.